Forty seven clinical samples of
Fowl adenovirus (FAdV) associated with Inclusion Body Hepatitis (IBH) from
Peruvian broilers received between July 2006 and April 2013 were genotyped
using sequencing of L1 Loop of Hexon gene. All 47 clinical samples presented
macroscopic and histopathology lesions consistent with IBH, and amplified a
specific fragment of Hexon gene by Polymerase Chain Reaction (PCR). A unique
nucleotide sequence of 789 base pairs of Hexon gene (position 273 to 1061) was
obtained in all 47 clinical samples analyzed. This sequence showed a high level
of conservation in amino acid and nucleotide sequence (>99%) with a Fowl
Adenovirus C serotype 4 previously identified. Sequence and phylogenetic
analysis indicate no genotypic variation in Peruvian isolates. The presence of
a unique genotype very closely related with genotype C1 previously reported in
Peru and Ecuador (>99%), suggests the presence of FAdV C serotype 4 genotype
C1 in clinical cases of IBH from Peruvian broilers.
Fowl Adenovirus C Molecular Characterization Inclusion Body Hepatitis Peru1. Introduction
Fowl Adenovirus (FAdV) includes 12 serotypes [1] , with a high diversity of strains for each serotype and wide range of clinical and pathological presentations (pneumonias, tracheitis, gizzard erosions, proventriculitis, pancreatitis, inclusion body hepatitis and hydropericardium) [2] . The most important sanitary problem in North America and South America is inclusion body hepatitis/hydropericardium syndrome due to high levels of morbidity and mortality in broilers resulting in significant economic losses [3] -[7] . This great diversity exhibited by FAdV required the use of additional techniques to make a correct identification of levels of species and serotypes and characterization of genetic diversity. An alternative to microbiologic and serologic techniques are molecular techniques based on DNA analysis. Several studies have reported the feasibility of molecular genetic tools for genetic differentiation and molecular typing of FAdV. The use of PCR for Hexon gene, RFLP analysis and direct sequencing of Hexon and Penton genes [8] -[10] have permitted a correct identification and characterization of clinical isolates. The lack of information available for molecular characterization of isolates from Peru do not allow the identification of the type of serotypes currently present in poultry production systems. This is a key requirement for establishing actions to control and prevent IBH. The objective of this study was to determine FAdV genotypes/serotypes isolated from IBH clinical cases in broilers from Peru.
2. Materials and Methods2.1. Sources of Samples
A total of 47 suspected clinical cases of Inclusion body hepatitis (IBH) from broilers received in the Microbiology Laboratory-Bioservice SRL between July 2006 and April 2013 were included in this study (Table 1). Agar gel immune-diffusion (AGID) tests for serotypes 4, 8 and 9, liver histopathology analysis and Hexon gene PCR were performed to confirm the presence of fowl adenovirus (FAdV) in all suspected clinical cases.
2.2. Viral DNA Extraction and PCR Amplification of Hexon Gene
Viral DNA was extracted from liver lysate using DNeasy Tissue kit (Qiagen) according to manufacturer. PCR for L1 loop of the Hexon gene was carried out using primers HA and HB as described previously [9] . In brief, the PCR reaction was performed using a final volume of 20 uL containing 10 ng genomic DNA, 1X PCR buffer (20 mM Tris-HCl, 20 mM KCl, 5 mM (NH4)2SO4), 2 mM MgCl2, 5 pmol of each primer, 0.2 mM of dNTPs, 0.5 U Maxima HotStart-Taq DNA polymerase (Fermentas). Thermal cycles were the following: initial denaturation of 94˚C for 3 minutes, 30 cycles of 94˚C for 30 seconds, 53˚C for 60 seconds, 72˚C for 30 seconds and a final extension of 72˚C for 5 minutes. The PCR amplification was carried out using a Veriti Thermal cycler (Applied Biosystems). The PCR products were separated in 1% agarose gel TBE 1X electrophoresis and visualized by fluorescence using an ethidium bromide solution (0.1 ug/mL) and UV light.
2.3. Sequencing and Phylogenetics Analysis of L1 Loop of Hexon Gene
The PCR products were purified using the Wizard® SV Gel and PCR Clean-Up kit, cloned with pGEM-T Easy (Promega) and submitted for sequencing to Macrogen Inc, Korea. The sequencing reaction was performed in both directions using Big Dye sequencing kit (Applied Biosystems) and the primers HA and HB. Partial nucleotide sequence of the L1 Loop of hexon gene from FAdV’s was submitted to Gene Bank with the following accession number: KF601685.
Forward and reverse sequences were aligned together using Clustal program [11] to create consensus se- quences. The consensus nucleotide sequences were aligned by the Clustal W method using MEGA 5.0 software [12] . Pairwise amino acid and nucleotide identity and pairwise evolutionary distances were computed using
Table 1
. Collection year and city of origin of 47 clinical cases of Inclusion body hepatitis (IBH) from broilers from Peru
Year of isolation
Place of isolation (City, Province)
Number of isolates
2006, 2007, 2011, 2012, 2013
Lima, Lima
12
2011, 2012, 2013
Huaral, Lima
20
2011, 2012, 2013
Chilca, Lima
1
2011
Iquitos, Loreto
2
2011
Arequipa, Arequipa
1
2011
Tacna, Tacna
1
2011
Chanchamayo, Junin
2
2011
Nazca, Ica
1
2006, 2007, 2012, 2013
Trujillo, La Libertad
7
Mega v5.0. A phylogenetic analysis for the distance method was performed using partial sequences of L1 loop of Hexon gene and Hexon protein previously published in Gene Bank and FAdV Peruvian sequence (Table 2)
Table 2
. ID sequence, accession number, country of origin, species, serotype and genetic cluster of 37 FAdV using phylogenetic analysis
ID Sequence
Accession number1
Country2
Species3
Serotype4
Genetic Cluster5
FAdV_1_(CELO)
AAL13217
NR6
A
EU/US 1
A
FAdV_2_(SR48)
AAN77072
NR6
D
EU 2
D2
FAdV_3_(75)
AAN77075
NR6
D
EU 3
D3
FAdV_4_(KR5)
AAN77077
NR6
C
EU 4
C
FAdV_5_(VR-830)
AAL13222
NR6
E
US 5
D1
FAdV_5_(TR22)
AAN77079
NR6
B
EU 5
E
FAdV_5_(340)
AAN77078
NR6
B
EU 5
B
FAdV_6_(VR-831)
AAL13224
NR6
D
US 6
D3
FAdV_6_(CR119)
AAN77080
NR6
E
EU 6
D1
FAdV_7_(YR36)
AAN77081
NR6
E
EU 7
D1
FAdV_7_(VR-832)
AAL13225
NR6
E
EU 7
D1
FAdV_8_(TR59)
AAN77082
NR6
E
EU 8
D1
FAdV_8_(VR-833)
AAL13221
NR6
D
US 8
D3
FAdV_9_(VR-834)
AAL13226
NR6
C
US 9
C
FAdV_9_(764)
AAN77084
NR6
E
EU 9
D1
FAdV_10_(VR-835)
AAL13227
NR6
E
US 10
D1
FAdV_11_(X11)
AAL13223
NR6
E
US 11
D1
FAdV_11_(C2B)
AAN77085
NR6
C
EU 11
D2
FAdV_12_(380)
AAL13228
NR6
D
US 12
D2
(922-1)_Germany
FN869978.1
Germany
C
EU/US 4
C
(09-2602)_Austria
FN869977.1
Austria
C
EU/US 4
C
(K31)_Pakistan
FN869976.1
Pakistan
C
EU/US 4
C
(09-584)_Austria
FN869975.1
Austria
C
EU/US 4
C
(53)_Peru
FN869973.1
Peru
C
EU/US 4
C
(K1013)_Ecuador
FN869972.1
Ecuador
C
EU/US 4
C
(Da60)_Germany
FN869971.1
Germany
C
EU/US 4
C
(K99-97)_Kuwait
FN869970.1
Kuwait
C
EU/US 4
C
(AG234)_Mexico
FN869969.1
Mexico
C
EU/US 4
C
South_Korea
HQ697593.1
South Korea
C
EU/US 4
C
(04-50388)_Canada
EF685395.1
Canada
C
EU/US 4
C
(488)_Russia
AY581295.1
Russia
C
EU/US 4
C
(4158)_Italy
HM592284.1
Italy
C
EU/US 4
C
(5997)_Italy
HM592281.1
Italy
C
EU/US 4
C
(6169)_Italy
HM592277.1
Italy
C
EU/US 4
C
(5670)_Italy
HM592274.1
Italy
C
EU/US 4
C
(488)_Indian
AY581295.1
India
C
EU/US 4
C
(HARYANA-07)_Indian
EU847626.1
India
C
EU/US 4
C
1Accesion number for Hexon protein or sequence in database Genbank or EMBL. 2Country of origin of strain. 3Fowl Adenovirus species according with [10] . 4Fowl Adenovirus serotype according with [1] . 5Fowl Adenovirus phylogenetic cluster according with [10] . 6No reported in [10] .
using Kimura-2-parameter method [13] and Dayhoff matrix based method [14] . A dendogram was constructed with the Neighbor-joining method [15] using 1000 bootstrap [16] replicate values with Mega v5.0.
3. Results3.1. Clinical Cases of Inclusion Body Hepatitis
All clinical cases presented liver histopathology consistent with IBH lesions hepatocyte cytoplasmic vacuolation, multifocal to coalescing areas of hepatocytes necrosis, irregular shape hepatocyte nuclei, presence of basophilic intranuclear inclusion bodies without presence of HPS. The presence of FAdV was confirmed by PCR amplification of Hexon gene fragment in 47 suspected clinical cases of IBH.
3.2. Sequencing and Phylogenetic Analysis of L1 Loop of Hexon Gene
All 47 Peruvian FAdV isolates did not show genotype variation, a unique nucleotide sequence of 789 bp corresponding to position 273 and 1061 of Hexon gene amplified showed high nucleotide identity level (99.5% identity, E value = 0) and high conservation level in the amino acidic sequence (99.5%) in relation with a FAdV C serotype 4 previously identified [10] . The pairwise matrix of evolutionary divergence (nucleotide and amino acid) showed less divergence between Peruvian FAdV with FAdV C serotype 4 (0.005, 0.005) and FAdV C serotype US 9 (0.029, 0.070) in contrast to FAdV D serotype 8 (0.365, 0.709) and serotype 12 (0.378 and 0.663) (Table 3). Phylogenetic analysis from a segment of 770 nucleotide (260 amino acids) suggests the presence of FAdV group C serotype 4 present in all clinical cases from IBH of Peruvian broilers (Figure 1(a) and
Figure 1
(a) Phylogenetic tree of L1 loop of the hexon nucleotide sequence from 12 species of FAdV’s. The evolutionary relationships of taxa were inferred using Neighbor-Joining method [17] . The evolutionary distances were computed using the Kimura 2-parameter method [13] . The analysis involved 20 nucleotide sequences and 770 nucleotide positions in the final dataset, the strain names are according to Table 2. Bootstrap [16] test (1000 replicates) is expressed in % values. The scale bar indicates the evolutionary distance between sequences; (b) Phylogenetic tree of L1 loop of the hexon protein sequence from 12 species of FAdV’s. The evolutionary relationships of taxa were inferred using Neighbor-Joining method [17] . The evolutionary distances were computed using the Dayhoff matrix method [14] . The analysis involved 20 amino acid sequences and 199 amino acid positions in the final dataset, the strain names are according to Table 2. Bootstrap [16] test (1000 replicates) is expressed in % values. The scale bar indicates the evolutionary distance between sequences
Table 3
. Pairwise estimates of evolutionary divergence1 of nucleotide sequence(below diagonal) and amino acid sequence (above diagonal) between 12 species of FAdV2
FAdV Peru
FAdV1
FAdV2
FAdV3
FAdV4
FAdV5A
FAdV5B
FAdV5C
FAdV6A
FAdV6B
FAdV7A
FAdV7B
FAdV8A
FAdV8B
FAdV9A
FAdV9B
FAdV10
FAdV11A
FAdV11B
FAdV12
FAdVPeru
0.628
0.663
0.704
0.005
0.603
0.663
0.573
0.704
0.568
0.578
0.573
0.548
0.709
0.070
0.603
0.598
0.583
0.663
0.663
FAdV1
0.300
0.573
0.608
0.628
0.583
0.648
0.598
0.608
0.603
0.553
0.563
0.578
0.603
0.618
0.583
0.598
0.553
0.563
0.563
FAdV2
0.374
0.335
0.432
0.663
0.558
0.663
0.563
0.432
0.508
0.543
0.528
0.528
0.427
0.658
0.558
0.553
0.538
0.030
0.030
FAdV3
0.366
0.323
0.203
0.704
0.568
0.588
0.588
0.000
0.588
0.563
0.558
0.578
0.101
0.724
0.568
0.573
0.563
0.417
0.417
FAdV4
0.005
0.299
0.374
0.365
0.603
0.663
0.573
0.704
0.568
0.578
0.573
0.548
0.709
0.070
0.603
0.598
0.583
0.663
0.663
FAdV5A
0.353
0.321
0.278
0.273
0.352
0.573
0.543
0.568
0.317
0.216
0.246
0.271
0.563
0.613
0.010
0.035
0.221
0.563
0.563
FAdV5B
0.361
0.340
0.303
0.264
0.360
0.278
0.533
0.588
0.573
0.558
0.568
0.543
0.598
0.653
0.573
0.568
0.558
0.653
0.653
FAdV5C
0.332
0.312
0.295
0.305
0.331
0.284
0.271
0.588
0.503
0.523
0.518
0.528
0.583
0.588
0.543
0.538
0.518
0.563
0.563
FAdV6A
0.369
0.326
0.205
0.003
0.368
0.275
0.266
0.308
0.588
0.563
0.558
0.578
0.101
0.724
0.568
0.573
0.563
0.417
0.417
FAdV6B
0.339
0.313
0.240
0.258
0.339
0.151
0.279
0.262
0.261
0.286
0.281
0.302
0.573
0.563
0.322
0.312
0.286
0.533
0.533
FAdV7A
0.336
0.301
0.262
0.268
0.336
0.096
0.278
0.265
0.270
0.117
0.055
0.231
0.568
0.578
0.221
0.211
0.010
0.553
0.553
FAdV7B
0.331
0.305
0.257
0.264
0.332
0.105
0.277
0.264
0.266
0.116
0.021
0.266
0.568
0.573
0.241
0.241
0.065
0.543
0.543
FAdV8A
0.339
0.310
0.249
0.274
0.339
0.145
0.270
0.268
0.277
0.140
0.122
0.135
0.573
0.543
0.271
0.256
0.236
0.533
0.533
FAdV8B
0.365
0.323
0.200
0.044
0.364
0.271
0.275
0.308
0.047
0.260
0.265
0.264
0.279
0.714
0.563
0.573
0.568
0.412
0.412
FAdV9A
0.029
0.301
0.371
0.368
0.031
0.353
0.355
0.334
0.370
0.340
0.339
0.336
0.338
0.364
0.613
0.613
0.583
0.668
0.668
FAdV9B
0.355
0.322
0.277
0.271
0.353
0.009
0.278
0.283
0.274
0.151
0.094
0.100
0.145
0.271
0.355
0.035
0.226
0.563
0.563
FAdV10
0.356
0.327
0.274
0.274
0.355
0.021
0.278
0.283
0.277
0.144
0.090
0.099
0.138
0.273
0.357
0.021
0.216
0.568
0.568
FAdV11A
0.338
0.300
0.260
0.266
0.338
0.097
0.277
0.262
0.269
0.117
0.003
0.023
0.123
0.264
0.340
0.095
0.091
0.548
0.548
FAdV11B
0.378
0.332
0.010
0.199
0.378
0.281
0.296
0.295
0.201
0.248
0.269
0.262
0.252
0.196
0.378
0.279
0.277
0.266
0.000
FAdV12
0.378
0.335
0.010
0.201
0.378
0.282
0.299
0.296
0.204
0.248
0.270
0.262
0.253
0.199
0.378
0.281
0.278
0.268
0.003
1Analyses were conducted using p-distance [18] . This distance is the proportion (p) of amino acid (nucleotide) sites at which the two sequences to be compared are different. 2The FAdV strain accession number: FAdV1 (AAL13217), FAdV2 (AAN77072), FAdV3 (AAN77075), FAdV4 (AAN77077), FAdV5A (AAL13222), FAdV5B (AAN77079), FAdV5C (AAN77078), FAdV6A (AAL13224), FAdV6B (AAN77080), FAdV7A (AAN77081), FAdV7B (AAL13225), FAdV8A (AAN77082), FAdV8B (AAL13221), FAdV9A (AAL13226), FAdV9B (AAN77084), FAdV10 (AAL13227), FAdV11A (AAL13223), FAdV11B (AAN77085), FAdV12 (AAL13228) and FAdVPeru (this study).
Figure 1(b)). The FAdV genotype identified is closely related with the genotype C1 [19] previously reported in Peru (evolutionary divergence = 0, 100% nucleotide identity) and Ecuador (evolutionary divergence = 0.002, 99.9% nucleotide identity), however the two isolates showed 100% amino acid identity with FAdV isolated in this study (Table 4); this relationship is corroborated by phylogenetic clustering analysis (Figure 2(a) and Figure 2(b)).
4. Discussion
The nucleic acid technology has demonstrated the utility to detect differences in structural proteins related with immune response as Hexon protein. The sequencing of Hexon gene permits a correct identification of species level and serotype in FAdV, with similar results as reported for RFLP genome analysis [2] . Inside Hexon gene, 7 hypervariable regions were identified exclusively located in Loop 1 (L1) and Loop 2 (L2) regions [20] . The loop 1 in Hexon gene of FAdV reported higher variability [8] , and has demonstrated additional applications than include genetic variability analysis of FAdV isolates [6] [10] [21] .
Figure 2
(a) Phylogenetic tree of L1 loop of the hexon nucleotide sequence from FAdV C serotype 4. The evolutionary relationships of taxa were inferred using Neighbor-Joining method [17] . The evolutionary distances were computed using the Kimura 2-parameter method [13] . The analysis involved 19 nucleotide sequences and 611 nucleotide positions in the final dataset, the strain names are according to Table 2. Bootstrap [16] test (1000 replicates) is expressed in % values. The scale bar indicates the evolutionary distance between sequences. (b) Phylogenetic tree of L1 loop of the hexon protein sequence from FAdV C serotype 4. The evolutionary relationships of taxa were inferred using Neighbor-Joining method [17] . The evolutionary distances were computed using the Dayhoff matrix method [14] . The analysis involved 19 amino acid sequences and 202 amino acid positions in the final dataset, the strain names are according to Table 2. Bootstrap [16] test (1000 replicates) is expressed in % values. The scale bar indicates the evolutionary distance between sequences
During this study we analyzed 47 FAdV isolates from IBH clinical cases, examined their Hexon gene loop 1 sequences and compared them with reference strains previously published sequences. Our results suggest the absence of genetic variability in FAdV C in Peru, similar to previous reports [19] ; however this study only has access to one Peruvian isolate. Similar results were reported from India [22] [23] and South Korea [24] for clinical cases of IBH associated with the presence of FAdV C serotype 4 (>99% clinical cases of IBH). However, FAdV C serotype 4 (strain K531) is also associated with hydropericardium syndrome outbreaks (HPS) from South Korea [24] [25] and Germany [26] in contrast with Peruvian FAdV C serotype 4 than only reports IBH clinical lesions. The amino acid sequence analysis shows a slight differentiation between isolates from Peru and South Korea (Table 4) and phylogenetic analysis corroborates the different genetic clustering (Figure 2(B)).
The presence of multiple FAdV species in clinical cases of IBH were reported in Canada [6] with a fixation of FAdV serotypes 8 (45%) and 11 (45%) and South Korea [27] with FAdV serotype 4 (60%) and 11 (40%), however South Korean FAdV serotype 4 showed clinical hydropericardium/ IBH lesions in contrast to the Canadian report [6] .
This study is the first report of genetic characterization of FAdV C isolates from Peru that includes samples from a coastal region (Ica, La Libertad and Lima regions) which concentrates 80% of poultry production systems in Peru and provides important information than can serve as starting point for further investigations related to pathogenicity, antigenic properties and specific vaccine development for prevention and control of IBH
Table 4
. Pairwise estimates of evolutionary divergence1 of nucleotide sequences (below diagonal) and amino acid sequence (above diagonal) between 19 strain of FAdV-C serotype 4
FAdV Peru
Germany A
Austria A
Pakistan
Austria B
Peru
Ecuador
Germany B
Kuwait
Mexico
South Korea
Canada
Russia A
Italy A
Italy B
Italy C
Italy D
Indian A
Indian B
FAdV Peru
0.050
0.045
0.035
0.045
0.000
0.000
0.010
0.005
0.005
0.025
0.010
0.035
0.030
0.045
0.010
0.025
0.035
0.040
Germany A
0.029
0.005
0.015
0.010
0.050
0.050
0.040
0.045
0.045
0.045
0.040
0.015
0.030
0.005
0.045
0.030
0.015
0.020
Austria A
0.023
0.008
0.010
0.005
0.045
0.045
0.035
0.040
0.040
0.040
0.035
0.010
0.025
0.000
0.040
0.025
0.010
0.015
Pakistan
0.023
0.008
0.007
0.010
0.035
0.035
0.035
0.030
0.030
0.050
0.035
0.000
0.020
0.010
0.035
0.020
0.000
0.005
Austria B
0.026
0.011
0.007
0.010
0.045
0.045
0.040
0.040
0.040
0.045
0.040
0.010
0.020
0.005
0.035
0.020
0.010
0.015
Peru
0.000
0.029
0.023
0.023
0.026
0.000
0.010
0.005
0.005
0.025
0.010
0.035
0.030
0.045
0.010
0.025
0.035
0.040
Ecuador
0.002
0.031
0.025
0.025
0.028
0.002
0.010
0.005
0.005
0.025
0.010
0.035
0.030
0.045
0.010
0.025
0.035
0.040
Germany B
0.007
0.028
0.020
0.023
0.023
0.007
0.008
0.005
0.005
0.015
0.000
0.035
0.025
0.035
0.005
0.020
0.035
0.040
Kuwait
0.005
0.029
0.021
0.021
0.025
0.005
0.007
0.002
0.000
0.020
0.005
0.030
0.025
0.040
0.005
0.020
0.030
0.035
Mexico
0.007
0.031
0.023
0.023
0.026
0.007
0.008
0.003
0.002
0.020
0.005
0.030
0.025
0.040
0.005
0.020
0.030
0.035
South Korea
0.016
0.028
0.026
0.026
0.026
0.016
0.018
0.010
0.011
0.013
0.015
0.050
0.040
0.040
0.020
0.035
0.050
0.054
Canada
0.008
0.029
0.021
0.025
0.025
0.008
0.010
0.002
0.003
0.005
0.011
0.035
0.025
0.035
0.005
0.020
0.035
0.040
Russia A
0.023
0.008
0.007
0.000
0.010
0.023
0.025
0.023
0.021
0.023
0.026
0.025
0.020
0.010
0.035
0.020
0.000
0.005
Italy A
0.025
0.020
0.018
0.015
0.015
0.025
0.026
0.021
0.023
0.025
0.021
0.023
0.015
0.025
0.020
0.010
0.020
0.025
Italy B
0.026
0.005
0.003
0.003
0.007
0.026
0.028
0.023
0.025
0.026
0.023
0.025
0.003
0.015
0.040
0.025
0.010
0.015
Italy C
0.011
0.026
0.021
0.021
0.025
0.011
0.013
0.005
0.007
0.008
0.011
0.007
0.021
0.016
0.021
0.015
0.035
0.040
Italy D
0.021
0.020
0.018
0.015
0.015
0.021
0.023
0.018
0.020
0.021
0.018
0.020
0.015
0.003
0.015
0.013
0.020
0.025
Indian A
0.023
0.008
0.007
0.000
0.010
0.023
0.025
0.023
0.021
0.023
0.026
0.025
0.000
0.015
0.003
0.021
0.015
0.005
Indian B
0.025
0.010
0.008
0.002
0.011
0.025
0.026
0.025
0.023
0.025
0.028
0.026
0.002
0.016
0.005
0.023
0.016
0.002
1Analyses were conducted using p-distance [18] . This distance is the proportion (p) of amino acid (nucleotide) sites at which the two sequences to be compared are different. 2The FAdV strain accession number: Germany A (FN869978.1), Austria A (FN869977.1), Pakistan (FN869976.1), Austria B (FN869975.1), Peru (FN869973.1), Ecuador (FN869972.1), Germany B (FN869971.1), Kuwait (FN869970.1), Mexico (FN869969.1), South Korea (HQ697593.1), Canada (EF685395.1), Russia A (AY581295.1), Italy A (HM592284.1), Italy B (HM592281.1), Italy C (HM592277.1), Italy D (HM592274.1), Indian A (AY581295.1), Indian B (EU847626.1) and Fad VPeru (this study).
associated with FAdV.
5. Conclusion
In conclusion, our results indicate the presence of FAdV C, serotype 4, genotype C1 circulating in clinical cases of IBH from Peruvian broilers.
Acknowledgements
This work was supported by Bioservice SRL. The authors wish to thanks to Dr. Karina Mendoza, Dr. Katherine Porturas, Dr. Stephane Lovon, Dr. Brenda Jara, Dr. Madeline Garcia, Dr. Heberht Uchuya and Dr. Jhonatan Castro for their cooperation in the collection and isolation of viral samples. We would like to also thank Dr. Manuel Moro for editorial comments and suggestions.
NOTES@endMarkP#wang#_title:ep!!!
*Corresponding author.
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